Mouse Langerhans Cells Differentially Express an Activated T Cell-Attracting CC Chemokine

Abstract

Epidermal Langerhans cells represent an immature population of dendritic cells, not yet able to prime naïve T cells. Following in vitro culture Langerhans cells mature into potent immunostimulatory cells. We constructed a representative cDNA library of in vitro matured murine Langerhans cells. Applying a differential screening procedure 112 differentially expressed cDNA clones were isolated. Thirty-six clones represented cDNA fragments of the same gene, identifying it to be the most actively expressed gene induced in maturing Langerhans cells. A full-length cDNA was sequenced completely. The open reading frame codes for a protein of 92 amino acids containing a leader peptide of 24 amino acids, yielding a mature protein of 7.8 kDa molecular weight. Database searches revealed 99.4% sequence identity on the nucleotide level to the recently described mouse CC chemokine ABCD-1, as well as 74% sequence identity to the human CC chemokine, the macrophage-derived chemokine/stimulated T cell chemotactic protein. Expression was analyzed by reverse transcriptase-polymerase chain reaction on a large panel of cell types. Unlike the macrophage-derived chemokine, expression was not detected in macrophages stimulated by various cytokines. Expression is restricted to cultured Langerhans cells, in vitro cultured dendritic cells, and lipopolysaccharide-activated B cells. Recombinant protein was expressed in the yeast Pichia pastoris and purified to homogeneity. Whereas no chemotactic activity was observed in chemotaxis assays for naïve T cells, B cells, cultured dendritic cells, and Langerhans cells, a strong chemoattractant activity was exerted on activated T cells. Thus, production of this chemokine by dendritic cells may be essential for the establishment and amplification of T cell responses.

Identification of a Novel, Dendritic Cell-associated Molecule, Dectin-1, by Subtractive cDNA Cloning

“…By the PCR-based differential display between "freshly isolated" immature Langerhans cells versus "cultured" mature Langerhans cells, Ross et al (66) have identified about 500 cDNA fragments whose expression was either up-or down-regulated during their maturation. By screening a cDNA library constructed from the cultured Langerhans cells with differential hybridization, they also identified that the expression of fascin, an actin-bundling protein, was markedly up-regulated during maturation (67). By searching the Human Genome Sciences nucleotide data bases with a consensus motif of C-type lectins, Bates et al (68) identified a new C-type lectin, termed DC immunoreceptor (DCIR), which was expressed by short term human DC cultures generated from CD34 ϩ progenitors as well as from CD14 ϩ monocytes.…”

Differential Regulation of CCL22 Gene Expression in Murine Dendritic Cells and B Cells

“…The major function of CCL22 appears to be the induction of chemotactic migration of activated T cells (7)(8)(9)15), predominantly of the Th2 type (16) and of CD4 ϩ CD25 ϩ regulatory T cells (17), through its specific receptor CCR4. However, another as yet unidentified receptor has also been implicated (18,19).…”

“…However, another as yet unidentified receptor has also been implicated (18,19). The murine CCL22 is expressed by activated B lymphocytes and DC (8,9). Murine epidermal Langerhans cells (LC), an exceptionally homogeneous population of immature DC, produce CCL22 mRNA only upon maturation (9), whereas other myeloid DC transcribe CCL22 constitutively (8).…”

“…The murine CCL22 is expressed by activated B lymphocytes and DC (8,9). Murine epidermal Langerhans cells (LC), an exceptionally homogeneous population of immature DC, produce CCL22 mRNA only upon maturation (9), whereas other myeloid DC transcribe CCL22 constitutively (8). Epidermal LC and other immature DC represent sentinel cells of the immune system.…”

Transcriptional targeting of dendritic cells for gene therapy using the promoter of the cytoskeletal protein fascin

“…In the preceeding work, we analyzed differential gene expression in maturing DC and identified several genes of functional importance, which are selectively expressed by mature DC and only very few other cell types. 7,8 For a detailed analysis, we isolated the promoter of one of these genes, encoding the actin-bundling protein fascin. This structural protein crosslinks actin filaments and was shown by us to be pivotal for dendrite formation in DC.…”